The induced pluripotent stem cells (iPS cells) can be produced by introducing reprogramming factors into somatic cells (K. Takahashi and S. Yamanaka, Cell 126 (4), 663, 2006; WO2007/069666). The iPS cells thus produced have been confirmed to possess pluripotency, for example, by the fact that iPS cells contribute to the germline of chimeric mice (K. Okita, et al., Nature 448 (7151), 313, 2007; M. Wernig, et al., Nature 448 (7151), 318, 2007). Since the iPS cells can be produced by using cells derived from a patient to be treated and then induced to differentiate into cells of intended tissues, they are expected in the field of regenerative medicine to serve as a transplantation material free from rejection.
However, in chimeric mice derived from the iPS cells thus produced and their offsprings, possibility that a tumor is formed due to reactivation of the introduced c-myc gene cannot be denied. Thus, in order to reduce their tumor formation ability, methods without using a retrovirus carrying c-Myc gene have been developed to produce iPS cells, although their efficiency of induction is relatively low (M. Nakagawa, et al., Nat Biotechnol 26 (1), 101, 2008; M. Wernig, et al., Cell Stem Cell 2 (1), 10, 2008; WO2008/118820; WO2009/057831). Nonetheless, in consideration of the high efficiency with using c-Myc, it is more desirable to develop a method for selecting iPS cells with low tumor formation ability while using a retrovirus carrying c-Myc.
Similarly, although various methods have been developed for practical application of producing iPS cells, such as a method of preparing iPS cells adaptable to production of chimeric mice without employing selection by a drug (A. Meissner, et al., Nat Biotechnol 25(10), 1177 2007) and a method of produce iPS cells using cells from various tissues (Aoi, T. et al., Science 321, 699-702, 2008), the iPS cells thus produced are not always suitable for transplantation.
Accordingly, it is expected to establish methods not only for producing iPS cells but also for selecting iPS cells with high safety.